In the treatment of tumors, autoimmune diseases, allergies and tissue rejection reactions, it is a disadvantage that the currently available medicaments, such as chemotherapeutic agents, corticosteroids and immunosuppressive agents, have a potential of side effects which is sometimes considerable, due to their relative non-specificity. It has been attempted to moderate this by various therapeutical concepts. Especially the use of immunotherapeutic agents is an approach, which resulted in an increase of the specificity of medicaments, especially in tumor treatment.
If the immunotherapeutic agent is an immunotoxin, then a monoclonal antibody (moAb) or an antibody fragment, which has a kinetic affinity for surface markers of tumor cells, is coupled with a cytotoxic reagent. If the immunotherapeutic agent is an anti-immunoconjugate for the treatment of autoimmune diseases, tissue rejection reactions or allergies, a structure relevant to pathogenesis or a fragment thereof is coupled to a toxin component. It has been found that immunotoxins can be characterized by a high immunogenicity in clinical use. This causes the formation of neutralizing antibodies in the patient, which inactivate the immunotoxin. Generally, a repeated and/or continuous administration of the therapeutic agents is unavoidable for long-term curative effects. This is particularly clear in the suppression of tissue rejection reactions after transplantations, or in the treatment of autoimmune diseases, due to the partly demonstrated genetically caused predisposition to a pathogenic autoimmune reaction.
The peptidic cell poisons which have been mostly used to date and are thus best characterized are the bacterial toxins diphtheria toxin (DT) (Beaumelle, B. et al. 1992; Chaudhary, V. et al. 1990; Kuzel, T. M. et al. 1993; LeMaistre, C. et al. 1998), Pseudomonas exotoxin A (PE) (Fitz Gerald, D. J. et al. 1988; Pai, L. H. and Pastan, I. 1998), and the plant-derived ricin-A (Engert, A. et al. 1997; Matthey, B. et al. 2000; O'Hare, M. et al. 1990; Schnell, R. et al. 2000; Thorpe, P. E. et al. 1988; Youle, R. J. and Neville, D. M. J. 1980). The mechanism of cytotoxic activity is the same in all of these toxins despite of their different evolutionary backgrounds. The catalytic domain inhibits protein biosynthesis by a modification of the elongation factor EF-2, which is important to translation, or of the ribosomes directly, so that EF-2 can no longer bind (Endo, Y. et al. 1987; Iglewski, B. H. and Kabat, D. 1975).
In most of the constructs employed to date, the systemic application of immunotoxins results in more or less strong side effects. In addition to the “vascular leak” syndrome (Baluna, R. and Vitetta, E. S. 1997; Schnell, R. et al. 1998; Vitetta, E. S. 2000), thrombocytopenia, hemolysis, renal insufficiency and sickness occur, depending on the construct employed and the applied dosage. Dose-dependent and reversible liver damage could also be observed (Battelli, M. G. et al. 1996; Grossbard, M. L. et al. 1993; Harkonen, S. et al. 1987). In addition to the documented side effects, the immunogenicity of the constructs employed to be observed in the use of the immunoconjugates or immunotoxins is the key problem of immunotherapy (Khazaeli, M. B. et al. 1994). This applies, in particular, to the humoral defense against the catalytic domains employed, such as ricin (HARA) (Grossbard, M. L. et al. 1998), PE (Kreitman, R. J. et al. 2000), or DT (LeMaistre, E. F. et al. 1992). Theoretically, all non-human structures can provoke an immune response. Thus, the repeated administration of immunotoxins and immunoconjugates is subject to limitations. A logical consequence of these problems is the development of human immunotoxins, now named human cytolytic fusion proteins (Rybak, S. et al. 1992).
WO 01/80880 A1 discloses the use of granzyme B as a human immunoprotease. The cytotoxic lymphocyte serine proteinase granzyme B induces apoptosis of abnormal cells by cleaving intracellular proteins at sites similar to those cleaved by caspases. However, granzyme B has a number of efficient natural inhibitors that prevent granzyme B-mediated apoptosis in certain cell types.
Therefore the availability of a human serine protease with improved apoptotic activity and reduced sensitivity towards activity-inhibiting substances would be highly advantageous.